Toll-like Receptors (TLRs) form a family of pattern recognition receptors which have a key role in modulating the innate immune response, they are also involved in tissue repair, maintenance of tissue integrity and tumorigenesis. Eleven Toll-like Receptors have been identified in humans to date. The members of the Toll-like Receptor family are highly conserved, with most mammalian species having between 10 to 15 Toll-like Receptors. Each Toll-like Receptor recognises specific pathogen-associated molecular signatures. Toll-like Receptor 2 (TLR2, CD282, TLR-2) can be activated by peptidoglycan, lipoproteins, lipoteichoic acid and endogenous ligands.
A number of monoclonal antibodies which have binding specificity for Toll-like Receptor 2 are known. WO 01/36488 discloses an antibody, designated as TL2.1, which is derived from a hybridoma cell line deposited in accordance with the Budapest Treaty at the European Collection of Cell Cultures (ECACC) under the accession number 99102832. This antibody antagonises the activation of Toll-like Receptor 2 expressed on human cells.
WO 2005/028509 discloses a murine monoclonal antibody, designated T2.5, which specifically inhibits the activation of mammalian TLR2. The T2.5 monoclonal antibody is shown to be cross-reactive to both human and murine forms of TLR2. This document further contains experimental data which suggests that the murine TL2.1 anti-TLR2 monoclonal antibody, as disclosed in WO 01/36488, is not cross-reactive to both human and murine forms of TLR2, as was stated in the description of that patent application. Rather, the TL2.1 antibody is shown in WO 2005/028509 as only binding to human Toll-like Receptor 2 and not murine Toll-like Receptor 2. The T2.5 monoclonal antibody of WO 2005/028509 was raised against the extracellular domain of TLR2, and therefore has binding specificity to an epitope in that area of Toll-like Receptor 2.
WO 2005/019431 discloses an antibody which has binding specificity to TLR2, which is designated 11G7. This murine antibody can be derived from hybridoma cell line 11G7 as deposited with the American Type Culture Centre (ATCC) under the designation PTA-5014. The 11G7 monoclonal antibody selectively binds to the extracellular domain of TLR2 and can block the induction of cytokine production by human peripheral blood mononuclear cells (PBMCs) stimulated with an agonist which activates a heterodimer formed between Toll-like Receptor 1 (TLR1) and TLR2. The 11G7 antibody does not inhibit cytokine production by PBMCs stimulated with an agonist which induces signalling through a heterodimer formed between Toll-like Receptor 6 (TLR6) and TLR2.
The use of rodent monoclonal antibodies, such as murine monoclonal antibodies, for in-vivo therapeutic applications has been shown to be associated with the generation of undesirable immune responses which are generated by the subject to whom the antibody is administered. Such immune responses can result in the production of antibodies which effectively neutralise the effectiveness of the therapeutic antibody. Such immune responses are typically referred to as human anti-mouse antibody (HAMA) responses. HAMA responses compromise the therapeutic effectiveness of the administered antibody in a number of ways, including impairing the ability of the therapeutic antibody to reach its binding target, this compromising the therapeutic effect of the antibody.
A number of approaches have been developed to address the issue of unwanted HAMA responses being raised against therapeutic antibodies which are administered to individuals. Typically, these approaches involve techniques which result in the replacement of certain components of the mouse antibody with equivalent portions derived from a human antibody. Such approaches can, for example, result in the production of chimeric antibodies which comprise murine variable regions joined to human-derived constant regions. Alternatively, a technique known as “CDR grafting” can be employed, wherein the complementarity determining regions (CDRs) from a murine antibody are grafted into a framework provided by regions of human antibody light and heavy chain variable domains. This results in the production of an antibody which retains the binding specificity of the murine antibody, but where the only non-human components are the grafted murine CDRs regions.
However, in both of these approaches, the therapeutic effectiveness of the resulting humanised antibody can be impaired. For example, the murine variable region component of a chimeric antibody can still provide the basis for a HAMA response to be mounted there against. Further, where CDR grafted humanised antibodies are produced, it has been observed that simple transplantation of the CDR regions often results in a reduced therapeutic efficacy of the antibody due to the binding affinity of the antibody being diminished.
The inventor has therefore identified the need to generate fully humanised monoclonal antibodies which have binding specificity for Toll-like Receptor 2, and which antagonise TLR2 function, but which are essentially non-immunogenic in humans. Following extensive experimentation, the inventor has produced a fully humanised monoclonal antibody which have binding specificity to human Toll-like Receptor 2, and which antagonises TLR2 function irrespective of whether Toll-like Receptor 2 forms a heterodimer with Toll-like Receptor 1 or Toll-like Receptor 6. This TLR2 antagonistic antibody is not produced from previously known chimeric or CDR-grafting techniques and therefore does not contain any murine amino acid residues. Furthermore, the antibody is shown to mediate TLR2 neutralisation without the need to bind to the CD32 cell surface antigen, this being a functional requirement of other known Toll-like Receptor 2 antagonistic antibodies. Furthermore, the fully humanised antibody of the invention is the first fully human TLR2 neutralising antibody to be known in the art. The antibody does not exhibit any T cell epitopes and therefore neutralising antibodies are not raised there against when administered to a subject. The anti-TLR2 antagonistic antibodies further exhibit a broad level of cross-reactivity to Toll-like Receptor 2 as expressed by a wide range of mammalian cells, with binding of the fully humanised antibody being surprisingly observed to bind to Toll-like Receptor 2 expressed on human, mouse and monkey cells.